Mechanism Of Arabidopsis Thaliana Drought Response Regulated By A Long Noncoding RNA9137

Drought,as one of natural disasters,not only affects the normal growth and development of plants,but also reduces crop production,which is a serious threat to the food security of China.In order to maintain both stable and sustained growth of agricultural production in our country,we should not only focus on the utilization efficiency of water resources,but also pay more attention to the breeding of drought-tolerant plants.Unveiling the inherent molecular mechanism of plant responses to drought stress can improve the predictability of breeding and greatly accelerate the breeding process.Recent researches have shown that long noncoding RNAs(lnc RNAs)play important roles in plant stress response,but their functions and regulatory mechanisms regarding drought response in plants are still unknown.A long noncoding RNA,lincRNA9137,which is involved in the regulation of drought response in Arabidopsis thalian,has been identified in our laboratory before.In this study,we explored both function and molecular mechanism of lincRNA9137 through applying genetic,biochemical and molecular biological approaches.The main results are presented as follow:1)The lincRNA9137 mutants shown decreased sensitivity to ABA treatment and reduced drought tolerance.Subsequently,we observed that water loss rates of mutants' leaves were significantly increased,which is caused by the increasing of stomatal aperture.This result suggests that lincRNA9137 regulates plant responses to drought through ABA-mediated stomatal aperture regulation pathway.2)Tissue-specific expression analysis shown that lincRNA9137 mainly expressed in roots of Arabidopsis thaliana.Linc RNA9137 was mainly enriched in the nucleus based on nuclear-cytoplasmic fractionation experiment,implying that it may regulate gene expression at the transcriptional level.3)Mutated lincRNA9137 decreased the expression level of its nearby gene JMJ29,which encodes a histone demethylase.In addition,the reduced drought resistance and increased stomatal aperture were observed in plants loss of JMJ29 function,indicating that JMJ29 is a downstream gene regulated by lincRNA9137.4)RBP82,a protein interacted with lincRNA9137,has been successfully identified by the yeast three-hybrid assay.Following RNA pull-down experiment proved the interaction between lincRNA9137 and RBP82.RBP82 encodes an AP2-like transcription factor,and it located in the nucleus depend on subcellular localization analysis.The dual luciferase reporter assay shown that RBP82 has transcriptional activation activity.Two loss of function mutants including rbp82-1and rbp82-2 of RBP82 were generated through CRISPR/Cas9 system.Phenotypic analysis shown that rbp82 mutants have reduced drought tolerance and increased stomatal aperture.Moreover,the expression levels of JMJ29 in rbp82 mutants were lower than that in wild-type.These results indicate that lincRNA9137 regulates plant responses to drought through interacting with RBP82 cooperatively to regulate the transcription of JMJ29.Our results not only diversify the mechanism of gene regulation directed by lnc RNA,but also improve the signal pathway of plant responses to drought stress,which provides a new theoretical basis and genetic resources for crop breeding.